Introduction: Levosimendan is an extensively investigated inodilator showing also cardioprotective and antiinflammatory effects. The aim of our study was to explore the influence of levosimendan on polymorphonuclear leucocytes (PMN), a main source of reactive oxygen species, in vitro and in patients with acute heart failure or septic myocardial depression. Methods: PMN isolated from healthy volunteers were incubated with levosimendan in vitro. After stimulation with N-formyl-Met-Leu-Phe (fMLP) or phorbol 12-myristate 13-acetate (PMA) respiratory burst was quantified using a fluorescent dye. Apoptosis and expression of cell adhesion molecules of PMN were measured by flow cytometry. For determination of in vivo effects patients with acute heart failure (n = 16) or septic cardiac failure (n = 9) receiving levosimendan treatment were enrolled consecutively. PMN were isolated to measure respiratory burst activity before treatment as well as one and two hours after initiation of levosimendan administration. Furthermore inflammatory, hemodynamic and renal function parameters were obtained. Results: In vitro, levosimendan suppressed respiratory burst activity in fMLP or PMA stimulated PMN in a dose dependent manner by 30 ± 11% (P < 0.001) at 100 ng/mL and by 27 ± 17% (P < 0.001) at 1000 ng/mL respectively. Markers of apoptosis and PMN cell adhesion molecule expression remained unaffected by levosimendan treatment.In vivo, levosimendan treatment for two hours resulted in a significant reduction of PMA stimulated oxidative burst by 45% (P < 0.01) and fMLP stimulated oxidative burst by 49% (P < 0.05) in patients with acute heart failure. In patients suffering from septic shock levosimendan treatment decreased oxidative burst activity in unstimulated, fMLP and PMA stimulated PMN by 48% (P < 0.05), 46% (P < 0.01) and 43% (P < 0.01) respectively. Conclusions: Levosimendan appears to exert distinct immunomodulatory effects by decreasing oxidative burst activity of PMN. This property might contribute to the previously described cardioprotective effects of the drug.